NDLI: Enhanced expression of human cDNA by phosphoglycerate kinase promoter-puromycin cassette in the mouse transthyretin locus

Content Provider	Springer Nature Link
Author	Li, Zhenghua Zhao, Gang Shen, Jingling Araki, Kimi Haruna, Kyoko Iue, Seiya Wang, Jun Yamamura, Ken ichi
Copyright Year	2010
Abstract	To produce a humanized mouse, it is critical to obtain a correct expression of a human gene/cDNA after insertion into a mouse locus. We previously generated a targeted allele in which the PGK-neo cassette, flanked by lox71 and loxP, was inserted into the first exon of the mouse endogenous transthyretin (Ttr) gene in ES cells. Using these ES cells, we showed that a human transthyretin (TTR) cDNA with the PGK-puro cassette can be efficiently inserted into this locus by Cre-mediated recombination, and that the human TTR cDNA was expressed in a tissue-specific manner under the control of the mouse endogenous Ttr promoter. To examine whether the PGK-puro cassette or IRES could affect the expression of human TTR cDNA, we generated four mouse lines using Cre and Flp-mediated recombination. The mouse line containing the PGK-puro cassette, but not IRES, exhibited quantitatively and temporally similar expression of human TTR cDNA. Removal of the PGK-puro cassette significantly downregulated the expression of the cDNA. The insertion of IRES sequence upstream of the human TTR cDNA resulted in decreased expression, even in the presence of the PGK-puro cassette. The mouse line containing IRES, but not PGK-puro, showed the lowest level of expression. These results suggest that the PGK-puro cassette is necessary to obtain the enhanced expression of a co-existing human cDNA in the mouse Ttr locus, even though the expression of co-existing cDNA was under the control of the mouse endogenous promoter.
Starting Page	191
Ending Page	200
Page Count	10
File Format	PDF
ISSN	09628819
Journal	Transgenic Research
Volume Number	20
Issue Number	1
e-ISSN	15739368
Language	English
Publisher	Springer Netherlands
Publisher Date	2010-04-06
Publisher Place	Dordrecht
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	Phosphoglycerate kinase promoter Internal ribosomal entry site Humanized mouse Cre-mediated recombination Transthyretin Human Genetics Plant Sciences Animal Genetics and Genomics Plant Genetics & Genomics Molecular Medicine
Content Type	Text
Resource Type	Article
Subject	Genetics Animal Science and Zoology Biotechnology Agronomy and Crop Science

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